过表达海兔属Ras同源物I(ARHI)增加人SW480结肠癌细胞凋亡

Overexpression of aplysia Ras homolog I(ARHI ) increases apoptosis in colon cancer SW480 cells

目的观察海兔属Ras同源物I(ARHI)对SW480人结肠癌细胞凋亡的影响。方法采用pc DNA3.1-FLAG-ARHI质粒转染和G418筛选法建立稳定表达ARHI的SW480细胞,采用反转录PCR验证ARHI mRNA和蛋白的表达。流式细胞术检测过表达ARHI对结肠癌细胞凋亡的影响;Western blot法检测蛋白激酶B(Akt)、磷酸化Akt(p-Akt)、p53和Bcl-2的蛋白水平。结果成功建立过表达ARHI的SW480细胞,过表达ARHI的SW480细胞凋亡率明显增加,Bcl-2和p-Akt蛋白水平明显降低、P53蛋白表达水平增加,而Akt蛋白水平无明显变化。结论过表达ARHI通过抑制Akt水平增加SW480人结肠癌细胞凋亡。

Objective To observe the effect of aplysia Ras homolog I（ARHI） gene on human colon cancer cell（SW480） apoptosis.Methods The pc DNA3.1-FLAG-ARHI plasmid was adopted to transfect the SW480 cell line,and G418 screening to obtain stably ARHI-transfected SW480 cells.Reverse transcription PCR and Western blotting were used to detect mRNA and protein expression of ARHI,respectively.The flow cytometry was applied to detect the effect of ARHI overexpression on colon cancer cell apoptosis.The levels of protein kinase B（Akt）,phosphorylated Akt（p-Akt）,p53 and Bcl-2 were determined by Western blotting.Results SW480 cells stably overexpressing ARHI were successfully established.Cell apoptosis rate in the cells significantly increased.The protein levels of Bcl-2 and p-Akt substantially decreased,whereas the protein expression level of P53 increased; Akt had no obvious changes.Conclusion The overexpression of ARHI promotes colon cancer SW480 cell apoptosis by inhibiting the level of Akt.